In susceptible individuals, asthma is a complex disease characterized by heightened responses to relatively innocuous antigens encountered via the respiratory tract. In a shift in thinking away from simple imbalances in Th2 and Th1 responses, there is increasing evidence for a protective role for regulatory T cells in allergic disease where they exhibit control over effector Th1 and Th2 cells. Naturally-occurring CD4+CD25+Foxp3+ regulatory cells (nTregs) isolated from the lungs of naive mice and transferred intratracheally into sensitized recipients prior to challenge suppress all aspects of lung allergic responses including airway hyperresponsiveness (AHR), airway eosinophilia, Th2 cytokine production, and goblet cell metaplasia. Conversely, depletion of these cells with anti-CD25 enhanced all of these responses. Our data indicate that the suppressive phenotype of nTregs is dependent on CD8- MHC I interactions and their production of IL-10 and TGF2. In the absence of CD8-MHC I interactions, the phenotype of the nTregs converts to one which enhances the development of lung allergic responses, and is associated with increased IL-13 production and decreased Foxp3 expression. By contrast, signaling through the glucocorticoid inducible tumor necrosis factor receptor (GITR) attenuates the suppressive phenotype. We will define the role of these nTregs in regulating mast cell-dependent and -independent AHR and whether they are responsible for the tolerant state induced by repeated allergen challenge. Using biochemical tools, genetic manipulation and in vitro and in vivo approaches, we will define the underlying mechanisms whereby CD8-MHC I interactions signal and maintain Foxp3 expression and the suppressive phenotype, and determine how signaling through GITR-GITR-ligand attenuates suppression, subverting and converting nTregs to an enhancing phenotype, characterized by Th2 cytokine production. In these approaches, we will delineate the counter-regulatory signals provided through Foxp3 and GITR and the interplay between these two defining events that appear to govern the fate (suppressive or enhancing phenotype) of nTregs. For the first time, these studies will identify the indispensable role and mechanism whereby Foxp3+nTregs control the development of lung allergic responses in sensitized hosts exposed to allergen challenge. Elucidation of the molecular basis for the functional activation of nTregs and the underlying mechanisms dictating nTreg-mediated suppression or nTreg conversion to an enhancing phenotype will form the basis for the control of their function in diseases such as asthma. PUBLIC HEALTH RELEVANCE: Asthma now afflicts more than 30 million Americans and despite the introduction of new therapies, morbidity and mortality continue to increase. If we are to impact this disease, a greater understanding of the mechanisms underlying asthma progression, delineation of the regulatory pathways, and identification of new strategies are required. This proposal addresses novel pathways that may regulate airway hyperresponsiveness and inflammation, with the potential of revealing novel and important therapeutic strategies.